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Mucinous epithelial ovarian cancer (mEOC) is a rare subtype of epithelial ovarian cancer, characterized by poor responses to standard platinum-based chemotherapy. Polo-like kinase 1 (PLK1) is a key regulator of mitosis and cell cycle progression and its inhibition has been recently identified as a target in mEOC. In this study, we aimed to identify further therapeutic targets in mEOC using a CRISPR/Cas9 library targeting 3015 genes, with and without treatment with onvansertib, a PLK1 inhibitor. We identified twelve genes associated with cell survival (ZC2HC1C, RPA2, KIN17, TUBG1, SMC2, CDC26, CDC42, HOXA9, TAF10, SENP1, MRPS31, and COPS2) and three genes (JUND, CARD9, and BCL2L2) in synthetic lethality with onvansertib treatment. We validated that SENP1 downregulation is important for the growth of mEOC cells through esiRNA interference and the use of a pharmacological inhibitor Momordin Ic. The downregulation of CARD9 and BCL2L2 combined with subtoxic doses of onvansertib interfered with mEOC cell growth. Interestingly, the combination of navitoclax, an inhibitor of BcL2 family members including BCL2L2, was synergistic in all four of the mEOC cell lines tested and substantially induced cell death through apoptosis. These data support the use of a combination of navitoclax and onvansertib as a new therapeutic strategy for mEOC. [ABSTRACT FROM AUTHOR]

Abstract Background Circular RNAs (circRNAs) are implicated in the development of oral squamous cell carcinoma (OSCC). The aim of current research is to elucidate the role and mechanism of circ_0011946 in the functional behaviors of OSCC cells. Methods Circ_0011946, microRNA (miR)-216a-5p, B cell lymphoma-2-like 2 protein (BCL2L2) abundances were exposed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) or western blot. Cell proliferation, migration, invasion and apoptosis were detected by MTT, colony formation assay, transwell, wound-healing and flow cytometry assays, respectively. Target correlation was tested by dual-luciferase reporter and RNA pull-down assays. An in vivo xenograft experiment was employed to investigate the function of circ_0011946 on tumor growth in vivo. Results Circ_0011946 and BCL2L2 levels were increased, while miR-216a-5p level was decreased in OSCC tissues and cells. Circ_0011946 knockdown impeded proliferation, migration, and invasion, but promoted apoptosis in OSCC cells. Circ_0011946 functioned as a sponge for miR-216a-5p, and BCL2L2 was targeted by miR-216a-5p. Besides, miR-216a-5p or BCL2L2 knockdown partly attenuated the inhibitory influences of circ_0011946 silence or miR-216a-5p overexpression on OSCC cell progression. Furthermore, circ_0011946 post-transcriptionally regulated BCL2L2 through sponging miR-216a-5p. Moreover, circ_0011946 knockdown constrained OSCC tumor growth in vivo. Conclusion Circ_0011946 silence repressed OSCC cell proliferation, migration, and invasion, but promoted apoptosis through the regulation of the miR-216a-5p/BCL2L2 axis.

Abstract Background X inactivation-specific transcript (XIST) is the long non-coding RNA (lncRNA) related to cancer, which is involved in the development and progression of various types of tumor. However, up to now, the exact role and molecular mechanism of XIST in the progression of ovarian cancer are not clear. We studied the function of XIST in ovarian cancer cells and clinical tumor specimens. Methods RT-qPCR was performed to detect the expression levels of miR-335 and BCL2L2 in ovarian cancer cells and tissues. MTT and transwell assays were carried out to detect cell proliferation, migration, and invasion abilities. Western blot was performed to analyze the expression level of BCL2L2. The interaction between miR-335 and XIST/BCL2L2 was confirmed using a luciferase reporter assay. Results The inhibition of XIST can inhibit the proliferation invasion and migration of human ovarian cancer cells. In addition, the miR-335/BCL2L2 axis was involved in the functions of XIST in ovarian cancer cells. These results suggested that XIST could regulate tumor proliferation and invasion and migration via modulating miR-335/BCL2L2. Conclusion XIST might be a carcinogenic lncRNA in ovarian cancer by regulating miR-335, and it can serve as a therapeutic target in human ovarian cancer.

Introduction: Cataracts caused by old age are one of the most frequent causes for blindness and poor vision worldwide. In this study, we aimed to clarify the possible role of rs1894720 polymorphism in the pathogenesis of age-related cataract.Material and Methods: Rs1894720 polymorphism genotype was detected by TaqMan. Bioinformatics analysis, luciferase assay, real-time PCR, western blot, and protein density analysis were conducted to establish the correlations between MIAT and miR-26b as well as between BCL2L2 and miR-26b. Flow cytometry and MTT assay were also performed to observe the effect of MIAT/miR-26b/BCL2L2 signalling pathway on the status of cell apoptosis and viability.Results: MIAT functioned as an endogenous competing RNA to sponge miR-26b. In addition, BCL2L2 was identified as a target of miR-26b. Therefore, the expression of miR-26b was obviously suppressed by MIAT or anti-miR-26b, while the mRNA and protein expression of BCL2L2 was up-regulated in the presence of MIAT or anti-miR-26b. Moreover, the positive effect of MIAT on BCL2L2 expression was exerted via inhibition of the expression of miR-26b. In addition, the cells transfected with MIAT or anti-miR-26b showed suppressed expression of caspase-3 and reduced apoptosis index but higher cell viability, indicating that MIAT could suppress cell apoptosis via inhibition of miR-26b expression. Furthermore, the subjects carrying the GT and TT genotypes of single-nucleotide polymorphism (SNP) rs1894720 were associated with a higher risk of age-related cataracts, as indicated by their odds ratio (OR) and p-values.Conclusions: Rs1894720 SNP could down-regulate the expression of MIAT, thus leading to reduced BCL2L2 expression and enhanced epithelial cell apoptosis in the lens, eventually increasing the incidence of age-related cataract. [ABSTRACT FROM AUTHOR]

Background: Circular RNAs (circRNAs) are implicated in the development of oral squamous cell carcinoma (OSCC). The aim of current research is to elucidate the role and mechanism of circ_0011946 in the functional behaviors of OSCC cells.Methods: Circ_0011946, microRNA (miR)-216a-5p, B cell lymphoma-2-like 2 protein (BCL2L2) abundances were exposed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) or western blot. Cell proliferation, migration, invasion and apoptosis were detected by MTT, colony formation assay, transwell, wound-healing and flow cytometry assays, respectively. Target correlation was tested by dual-luciferase reporter and RNA pull-down assays. An in vivo xenograft experiment was employed to investigate the function of circ_0011946 on tumor growth in vivo.Results: Circ_0011946 and BCL2L2 levels were increased, while miR-216a-5p level was decreased in OSCC tissues and cells. Circ_0011946 knockdown impeded proliferation, migration, and invasion, but promoted apoptosis in OSCC cells. Circ_0011946 functioned as a sponge for miR-216a-5p, and BCL2L2 was targeted by miR-216a-5p. Besides, miR-216a-5p or BCL2L2 knockdown partly attenuated the inhibitory influences of circ_0011946 silence or miR-216a-5p overexpression on OSCC cell progression. Furthermore, circ_0011946 post-transcriptionally regulated BCL2L2 through sponging miR-216a-5p. Moreover, circ_0011946 knockdown constrained OSCC tumor growth in vivo.Conclusion: Circ_0011946 silence repressed OSCC cell proliferation, migration, and invasion, but promoted apoptosis through the regulation of the miR-216a-5p/BCL2L2 axis. [ABSTRACT FROM AUTHOR]

Background: X inactivation-specific transcript (XIST) is the long non-coding RNA (lncRNA) related to cancer, which is involved in the development and progression of various types of tumor. However, up to now, the exact role and molecular mechanism of XIST in the progression of ovarian cancer are not clear. We studied the function of XIST in ovarian cancer cells and clinical tumor specimens. Methods: RT-qPCR was performed to detect the expression levels of miR-335 and BCL2L2 in ovarian cancer cells and tissues. MTT and transwell assays were carried out to detect cell proliferation, migration, and invasion abilities. Western blot was performed to analyze the expression level of BCL2L2. The interaction between miR-335 and XIST/BCL2L2 was confirmed using a luciferase reporter assay. Results: The inhibition of XIST can inhibit the proliferation invasion and migration of human ovarian cancer cells. In addition, the miR-335/BCL2L2 axis was involved in the functions of XIST in ovarian cancer cells. These results suggested that XIST could regulate tumor proliferation and invasion and migration via modulating miR-335/BCL2L2. Conclusion: XIST might be a carcinogenic lncRNA in ovarian cancer by regulating miR-335, and it can serve as a therapeutic target in human ovarian cancer. [ABSTRACT FROM AUTHOR]

miRNAs have been pointed to play critical role in the development of congenital heart disease (CHD). miRNA-375-3p (miR-375-3p) was involved in cardiac dysfunction and cardiogenesis. However, no prior study had established a therapeutic role of miR-375-3p in CHD. We intended to investigate the effect and mechanism of miR-375-3p on apoptosis in hypoxic cardiomyocytes in vitro. Expression of miR-375-3p, forkhead box P1 (FOXP1) and Bcl2 like protein 2 (Bcl2l2) was detected using real-time quantitative PCR and western blot. Apoptosis was measured with MTT assay, flow cytometry and caspase-3 activity assay. The potential target binding between miR-375-3p and FOXP1/Bcl2l2 was predicted on DianaTools, and was validated by luciferase reporter assay and RNA pull-down assay. As a result, miR-375-3p was upregulated and FOXP1/Bcl2l2 was downregulated in maternal serum of women with fetal CHD and hypoxia-induced rat cardiomyocyte h9c2 cells. Hypoxia induced apoptosis rate elevation, caspase-3 activity promotion and viability inhibition in h9c2 cells; overexpression of miR-375-3p promoted, whereas knockdown of miR-375-3p antagonized hypoxia-induced effects in h9c2 cells. In addition, miR-375-3p was validated to negatively regulate FOXP1 and Bcl2l2 expression through target binding, and silencing of FOXP1 and Bcl2l2 could independently abate the anti-apoptosis role of miR-375-3p knockdown in hypoxic h9c2 cells. Collectively, blocking miR-375-3p suppressed hypoxia-evoked apoptosis of cardiomyocytes by targeting and upregulating FOXP1 and Bcl2l2. Our results might suggest maternal serum miR-375-3p as a potential biomarker for prenatal detection of fetal CHD. [ABSTRACT FROM AUTHOR]

Background/Aims: Long non-coding RNAs (lncRNAs) are theorized to play key roles in the development of heart diseases. However, the role of lncRNAs in cardiomyocyte apoptosis is largely unknown. The present study examined the role of lncRNA SNHG1 in the human cardiomyocytes (HCMs) apoptosis and explored the underlying molecular mechanisms. Methods: SNHG1, miR-195 and mRNA expression was detected by qRT-PCR; protein level was determined by western blot; cell viability was detected by MTT assay; cell apoptosis was evaluated by flow cytometry and caspase-3 activity assay; the interaction between SNHG1 and miR195 was examined by using luciferase reporter assay. Results: Hydrogen peroxide (H2O2) treatment significantly suppressed cell viability and increased cell apoptotic rate and caspase-3 activity in HCMs. Overexpression of SNHG1 attenuated the effects of H2O2 on HCMs viability and apoptosis; while SNHG1 exerted the opposite effects. SNHG1 was found to sponge miR-195 and suppress the expression of miR-195 in HCMs. Overexpression of miR-195 suppressed cell viability and induced apoptosis in HCMs, and miR-195 was found to negatively regulate the expression of BCL-2 like protein 2 (BCL2L2) via targeting its 3’ untranslated region. Overexpression of BCL2L2 partially reversed the effects of miR-195 overexpression on cell viability and cell apoptosis of HCMs. MiR-195 overexpression or BCL2L2 knockdown attenuated the effects of SNHG1 overexpression on cell viability, cell apoptosis and protein levels of cleaved caspase-3, cleaved caspase-9 and Bax in H2O2-treated HCMs. Conclusion: Our results suggest a novel SNHG1/miR-195/BCL2L2 axis in the regulation of cardiomyocyte apoptosis. Modulation of SNHG1 may represent a novel strategy to treat cardiomyocyte apoptosis-related heart diseases.

Abstract Objective Our study was designed to explore the association miR‐335‐5p and BCL2L2 and to investigate the influence of miR‐335‐5p/BCL2L2 axis on cisplatin‐resistant ovarian cancer cells. Methods Microarray analysis was used to determine differentially expressed microRNAs in primary and cisplatin‐resistant A2780 cells. Cell function experiments were conducted to investigate the effect of miR‐335‐5p on the cisplatin sensitivity of A2780 cells. The targeted relationship between BCL2L2 mRNA and miR‐335‐5p was validated through luciferase assay. Tumor xenograft was performed to confirm the function of miR‐335‐5p in restoring the cisplatin sensitivity of the ovarian cancer cells. Results MiR‐335‐5p was lowly expressed in cisplatin‐resistant A2780 cells. Overexpression of miR‐335‐5p reduced cell survival and enhanced cisplatin‐induced cell apoptosis. BCL2L2 mRNA was a target of miR‐335‐5p, and silencing of BCL2L2 showed the similar results on the cell viability as miR‐335‐5p overexpression. Conclusion Upregulation of miR‐335‐5p expression enhanced the cisplatin sensitivity of ovarian cancer cells through suppressing BCL2L2, suggesting the potential of miR‐335‐5p/BCL2L2 axis as a therapeutic target for the cisplatin resistance of patients with ovarian cancer.

Recurrent spontaneous abortion (RSA) is a common health problem that affects 1–5% of women in reproductive age. Plenty of studies have indicated that microRNAs (miRNAs) are involved in the occurrence of miscarriage. MiR-93 has a wide range of functions in mammalian tissues and plays an important role in many diseases especially for cancers. However, it remains unknown whether miR-93 is associated with human RSA. In this report, clinical samples revealed that miR-93 expression was significantly elevated in the villi tissues of RSA patients. Upregulation of miR-93 inhibited human trophoblast cells HTR-8/SVneo cell proliferation, migration, and invasiveness, but promoted cell apoptosis in vitro. Conversely, the downregulation of miR-93 reversed these effects. Bcl-2 like protein 2 (BCL2L2), a potential target gene of miR-93, was inversely correlated with miR-93 expression in the villi of clinical samples. Furthermore, the luciferase reporter system demonstrated that miR-93 directly downregulated the expression of BCL2L2 by binding a specific sequence of its 3′-untranslated region (3′UTR). Collectively, these data strongly suggest that miR-93 regulates trophoblast cell proliferation, migration, invasive, and apoptosis by targeting BCL2L2 expression and is involved in the pathogenesis of RSA. [ABSTRACT FROM AUTHOR]

Aberrant expressions of microRNAs have been reported to be strongly associated with the progression and prognosis of various tumors, including oral squamous cell carcinoma (OSCC). Recent studies on miRNA expression profiling have suggested that microRNA‐16 (miR‐16) may be dysregulated in OSCC. However, the tumorigenic roles and mechanisms of miR‐16 in OSCC are still largely unknown. In this study, we demonstrated that miR‐16 was specifically downregulated in both OSCC patients and cancer cell lines. In addition, functional roles of miR‐16 in vitro suggested that the miR‐16 mimic inhibited cell proliferation and induced apoptosis, whereas miR‐16 inhibitor displayed the opposite effects. Luciferase reporter assay and correlation analysis showed that AKT3 and BCL2L2 were directly targeted by miR‐16 and were inversely expressed with miR‐16 in OSCC. Moreover, restoration of AKT3 and BCL2L2 expression could partially reverse the cell proliferation inhibition and apoptosis induction caused by miR‐16. In xenograft nude mice, miR‐16 mimics decreased the expression of AKT3 and BCL2L2 and reduced the tumors volumes and weights, whereas the miR‐16 inhibitor exhibited adverse effects in the derived xenografts. In conclusion, the findings suggested that miR‐16 functions as a tumor suppressor miRNA to inhibit cell proliferation and induce apoptosis in OSCC through decreasing the oncogenes AKT3 and BCL2L2 and that miR‐16 could be a potential therapeutic target for OSCC. microRNA‐16 is downregulated in oral squamous cell carcinoma (OSCC) tissues and cell lines, it functions as a tumor suppressor microRNAs to inhibit cell proliferation and induce apoptosis in OSCC through decreasing the oncogenes AKT3 and BCL2L2. [ABSTRACT FROM AUTHOR]

Background/Aims: Long non-coding RNAs (lncRNAs) are theorized to play key roles in the development of heart diseases. However, the role of lncRNAs in cardiomyocyte apoptosis is largely unknown. The present study examined the role of lncRNA SNHG1 in the human cardiomyocytes (HCMs) apoptosis and explored the underlying molecular mechanisms. Methods: SNHG1, miR-195 and mRNA expression was detected by qRT-PCR; protein level was determined by western blot; cell viability was detected by MTT assay; cell apoptosis was evaluated by flow cytometry and caspase-3 activity assay; the interaction between SNHG1 and miR195 was examined by using luciferase reporter assay. Results: Hydrogen peroxide (H2O2) treatment significantly suppressed cell viability and increased cell apoptotic rate and caspase-3 activity in HCMs. Overexpression of SNHG1 attenuated the effects of H2O2 on HCMs viability and apoptosis; while SNHG1 exerted the opposite effects. SNHG1 was found to sponge miR-195 and suppress the expression of miR-195 in HCMs. Overexpression of miR-195 suppressed cell viability and induced apoptosis in HCMs, and miR-195 was found to negatively regulate the expression of BCL-2 like protein 2 (BCL2L2) via targeting its 3' untranslated region. Overexpression of BCL2L2 partially reversed the effects of miR-195 overexpression on cell viability and cell apoptosis of HCMs. MiR-195 overexpression or BCL2L2 knockdown attenuated the effects of SNHG1 overexpression on cell viability, cell apoptosis and protein levels of cleaved caspase-3, cleaved caspase-9 and Bax in H2O2-treated HCMs. Conclusion: Our results suggest a novel SNHG1/miR-195/BCL2L2 axis in the regulation of cardiomyocyte apoptosis. Modulation of SNHG1 may represent a novel strategy to treat cardiomyocyte apoptosis-related heart diseases. [ABSTRACT FROM AUTHOR]

Abstract: Objective: Our study was designed to explore the association miR‐335‐5p and BCL2L2 and to investigate the influence of miR‐335‐5p/BCL2L2 axis on cisplatin‐resistant ovarian cancer cells. Methods: Microarray analysis was used to determine differentially expressed microRNAs in primary and cisplatin‐resistant A2780 cells. Cell function experiments were conducted to investigate the effect of miR‐335‐5p on the cisplatin sensitivity of A2780 cells. The targeted relationship between BCL2L2 mRNA and miR‐335‐5p was validated through luciferase assay. Tumor xenograft was performed to confirm the function of miR‐335‐5p in restoring the cisplatin sensitivity of the ovarian cancer cells. Results: MiR‐335‐5p was lowly expressed in cisplatin‐resistant A2780 cells. Overexpression of miR‐335‐5p reduced cell survival and enhanced cisplatin‐induced cell apoptosis. BCL2L2 mRNA was a target of miR‐335‐5p, and silencing of BCL2L2 showed the similar results on the cell viability as miR‐335‐5p overexpression. Conclusion: Upregulation of miR‐335‐5p expression enhanced the cisplatin sensitivity of ovarian cancer cells through suppressing BCL2L2, suggesting the potential of miR‐335‐5p/BCL2L2 axis as a therapeutic target for the cisplatin resistance of patients with ovarian cancer. [ABSTRACT FROM AUTHOR]

The impact of N6-methyladenosine (m6A) modification on pri-miRNA in sepsis-induced cardiomyopathy (SICM), and its underlying regulatory mechanism, have not been fully elucidated. We successfully constructed a SICM mice model through cecal ligation and puncture (CLP). In vitro, a lipopolysaccharide (LPS)-induced HL-1 cells model was also established. The results showed that sepsis frequently resulted in excessive inflammatory response concomitant with impaired myocardial function in mice exposed to CLP, as indicated by decreases in ejection fraction (EF), fraction shortening (FS), and left ventricular end diastolic diameters (LVDd). miR-193a was enriched in CLP mice heart and in LPS-treated HL-1 cells, while overexpression of miR-193a significantly increased the expression levels of cytokines. Sepsis-induced enrichment of miR-193a significantly inhibited cardiomyocytes proliferation and enhanced apoptosis, while this was reversed by miR-193a knockdown. Furthermore, under our experimental conditions, enrichment of miR-193a in SICM could be considered excessively maturated on pri-miR-193a by enhanced m6A modification. This modification was catalyzed by sepsis-induced overexpression of methyltransferase-like 3 (METTL3). Moreover, mature miRNA-193a bound to a predictive sequence within 3′UTRs of a downstream target, BCL2L2, which was further validated by the observation that the BCL2L2-3′UTR mutant failed to decrease luciferase activity when co-transfected with miRNA-193a. The interaction between miRNA-193a and BCL2L2 resulted in BCL2L2 downregulation, subsequently activating the caspase-3 apoptotic pathway. In conclusion, sepsis-induced miR-193a enrichment via m6A modification plays an essential regulatory role in cardiomyocyte apoptosis and inflammatory response in SICM. The detrimental axis of METTL3/m6A/miR-193a/BCL2L2 is implicated in the development of SICM. [ABSTRACT FROM AUTHOR]

Cervical cancer is one of the most common cancers in females, with a high incidence and mortality around the world. However, the pathogenesis in cervical cancer is not completely known. In the present study, we investigated the role of miR-126-5p and Bcl2l2 in cervical cancer cells. First, miR-126-5p expression was aberrantly downregulated in human cervical cancer tumor tissues in comparison with normal tissues, as evaluated by RT-PCR. Consistently, the levels of miR-126-5p were also significantly reduced in cervical cancer cell lines when compared to normal cervical epithelial cells. Flow cytometric analysis showed that the rate of apoptosis of cervical cancer cells was significantly increased by miR-126-5p overexpression but inhibited by miR-126-5p inhibitor. A similar change pattern was observed in the expression of apoptosis-regulated protein caspase 3 in cervical cancer cells transfected with miR-126-5p mimic or inhibitor. By bioinformatic prediction with online databases and verification using luciferase reporter assay, we then identified that Bcl2l2 is a direct target of miR-126-5p in cervical cancer cells. The expression of Bcl2l2 was strongly downregulated by the miR-126-5p mimic but upregulated by the miR-126-5p inhibitor in cervical cancer cells, and Bcl2l2 expression was significantly increased in human cervical cancer tumor tissues, which was negatively correlated with miR-126-5p levels. Furthermore, we confirmed that the rate of apoptosis was significantly increased by Bcl2l2 silencing in cervical cancer cells, which was not affected by the miR-126-5p inhibitor. In addition, the increased apoptosis of cells by the miR-126-5p mimic was inhibited by Bcl2l2 overexpression. In summary, miR-126-5p plays an inhibitory role in human cervical cancer progression, regulating the apoptosis of cancer cells via directly targeting Bcl2l2. This might provide a potential therapeutic target for cervical cancer. [ABSTRACT FROM AUTHOR]

• Breast cancers display no alterations in anti-apoptotic BCL2 family members, besides amplifications in MCL1, which is more prevalent in basal cancers. • Subtypes of breast cancer are heterogeneous regarding the expression and reliance on antiapoptotic BCL2 proteins. • Levels of mRNA expression of anti-apoptotic family proteins do not significantly affect survival in breast cancer. Progress in therapies and improved outcomes in recent decades have followed a better understanding of breast cancers pathogenesis and their heterogeneity but new treatments are needed especially for metastatic disease which remains incurable. Inhibition of apoptosis is a hallmark characteristic of cancer and can be targeted for therapy. The five anti-apoptotic members of the BCL2 family are at the core of apoptosis execution and are involved in apoptosis evasion of transformed cells. Genetic lesions as well as mRNA regulation of these members in breast cancer and its sub-types and implications for survival outcomes were investigated using data from various publicly available databases. Genes encoding for anti-apoptotic BCL2 proteins are rarely mutated in breast cancer and copy number alterations are observed only in MCL1 gene which is amplified in a minority of breast cancer ranging from 1.6% to 18.7% in breast cancers. Over-expression of BCL2, BCL-X and MCL1 is observed in luminal A cancers, while cases of luminal B and basal breast cancers display mRNA up-regulation of BCL-X and MCL1, respectively. Basal cancers possess also more frequently than other sub-sets MCL1 amplifications. Survival outcomes are not significantly different in cancers with higher expression of anti-apoptotic BCL2 mRNAs. Therapeutic targeting of the apoptotic process in breast cancer sub-types will be improved by a detailed understanding of the core players in the process, including anti-apoptotic BCL2 family proteins. A sub-set of breast cancers harbor amplifications of MCL1 and dysregulations of expression of most family members that could affect the sensitivity to their inhibition by altering the cell's apoptotic threshold. [ABSTRACT FROM AUTHOR]

• BCL2L2 and nine alternative splicing transcripts were cloned and identified in pigs. • One SP1 and two C/EBPα binding sites were characterized in the promoter. • Alternative splicing in 5′ UTR regulates the expression of BCL2L2. • Upstream opening reading frame represses the expression of BCL2L2. B cell lymphoma-2-like 2 (BCL2L2), an important regulator of apoptosis, plays vital roles in several physiological processes, as revealed by studies in humans and mice. However, reports on pig BCL2L2 are few, and the encoding gene has not been identified experimentally. This study was designed to clone the porcine BCL2L2 gene and its alternative splicing (AS) transcripts using molecular biological techniques and to analyze the regulatory mechanisms underlying transcription and translation. The BCL2L2 cDNA (V1) was 807 bp in length and encoded a polypeptide of 193 aa containing four BCL-2 homology domains. A total of nine AS transcripts were obtained, among which V2 and V3 differed from V1 in the 5′ untranslated region (UTR). The core promoter was mapped to a range of −1102 to −759 bp (the first nucleotide of the start codon was designated as +1). There were several functional cis -elements, including one SP1 and two C/EBPα binding sites at around −759 bp. AS in the 5′ UTR is involved in the regulation of gene expression, as revealed by dual-luciferase reporter and western blot analysis, and the secondary structure of the 5′ UTRs may be the reason for the differential expression of V1–3. At the same time, an upstream open reading frame (ORF) existed in each of the three 5′ UTRs, was found to repress the expression of the main ORF. Additionally, the roles of porcine BCL2L2 in cell proliferation and apoptosis were preliminarily analyzed. The results will contribute to further characterizing the role of BCL2L2. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) play a critical role in cancer development and progression. Aberrant expression of miR-15a has recently been reported in several cancers, but its role in non-small cell lung cancer (NSCLC) still remains obscure. We investigated the effects of miR-15a on proliferation, apoptosis, and metastasis in A549 cells. Eighteen paired NSCLC and adjacent non-tumor lung tissues were surgically removed and immediately snap frozen until total RNA was extracted and confirmed by two independent pathologists. The targets of miR-15a were predicted by bioinformatics tools. RNA isolation and quantitative real-time PCR (qRT-PCR), Western blot analysis, cell proliferation assay, cell cycle analysis, cell apoptosis assay, and migration and invasion assays were done. The wild type (WT) or mutant type (MT) 3′-untranslated region (UTR) vectors were co-transfected with miR-15a or negative control into A549 cells, and after 24 h of transfection, luciferase activity was measured using the Dual-Glo luciferase assay kit. Statistical analysis was performed using SPSS 13.0 software (SPSS, Chicago, IL, USA). P values of less than 0.05 were considered statistically significant. miR-15a was significantly downregulated in NSCLC than in adjacent non-cancerous tissues. miR-15a overexpression remarkably inhibited cell viability, invasion, and migration and promoted the apoptosis of NSCLC cells. Additionally, inhibition of miR-15a expression had the opposite effects on tumor progression, while cell cycle remained unaltered. Furthermore, we identified that BCL2L2 was a target of miR-15a and negatively regulated by miR-15a at the translational level. miR-15a acts as a tumor suppressor in NSCLC by directly targeting BCL2L2 and may serve as a potential diagnostic biomarker and therapeutic target for NSCLC. [ABSTRACT FROM AUTHOR]

Abstract: MiR-214 has been shown to inhibit cell growth, migration and invasion. Here we demonstrate that ectopic expression of miR-214 reduces cell survival, induces apoptosis and enhances sensitivity to cisplatin through directly inhibiting Bcl2l2 expression in cervical cancer HeLa and C-33A cells. Further analysis reveals that apoptosis induced by miR-214 is correlated with increased expression of Bax, caspase-9, caspase-8 and caspase-3. Moreover, we show that miR-214 is regulated by DNA methylation and histone deacetylation. Taken together, these data suggest that miR-214 might be a candidate target for the development of novel therapeutic strategies to treat cervical cancer. [Copyright &y& Elsevier]

Abstract: Influenza A virus is a cytolytic virus that induces apoptosis in numerous cell types, which contributes to cellular and organ dysfunction. MicroRNAs (miRNAs) represent a family of small noncoding RNAs controlling tanslation and transcription of many genes. Recent studies have revealed that miR-29c is involved in a variety of biological processes, including apoptosis. However, its role in influenza A virus infection is not well understood. Here, we report that miR-29c is involved in apoptosis induced by influenza A virus infection. We found that several apoptosis-associated miRNAs were stimulated in influenza A virus-infected A549 cells by miRNA array analysis. Within those, miR-29c was significantly up-regulated. In silico target prediction analysis revealed complementarity of miR-29c to the 3′-untranslated region (UTR) of BCL2L2 mRNA. Targeting of BCL2L2 3′ UTR by miR-29c was determined by luciferase assay. Functional overexpression of miR-29c with miR-29c precursor inhibited BCL2L2 protein expression. Transfection of miR-29c inhibitor abolished both suppression of BCL2L2 protein expression and A549 cells apoptosis induced by influenza A virus. Moreover, BCL2L2 overexpression rescued A549 cell death induced by influenza A virus infection. These findings indicate that miR-29c-mediated BCL2L2 suppression is involved in influenza virus-induced cell death in A549 cells. [Copyright &y& Elsevier]

It is increasingly clear that microRNAs (miRNAs) play an important role in controlling cell survival. However, the functional significance of miRNAs in ischemic brain injury remains poorly understood. In the present study, we assayed the expression levels of miR-29b after ischemic brain injury, and defined the target genes and biological functions of miR-29b. We found that the miR-29b levels were significantly increased in rat brain after transient middle cerebral artery occlusion and neurons after oxygen-glucose deprivation. Moreover, ectopic expression of miR-29b promoted neuronal cell death, whereas its repression decreased cell death. Furthermore, we verified that miR-29b directly targeted and inhibited Bcl2L2 gene expression, and then increased neuronal cell death. Importantly, Bcl2L2 overexpression rescued neuronal cell death induced by miR-29b. These results suggest an important role of miR-29b in regulating neuronal cell death, thus offering a new target for the development of therapeutic agents against ischemic brain injury. [ABSTRACT FROM AUTHOR]

Abstract: This study aimed to investigate the effects of fisetin, a common dietary natural flavonoid, on apoptosis of Huh-7 cells. The MTT assay was used to evaluate the reduction of cell viability. In the DNA fragmentation assay and comet assay, fisetin-induced DNA damage was visible as a formation of DNA fragmentation and comet tails. Fisetin also induced intracellular accumulation of reactive oxygen species. Two-dimensional gel electrophoresis was performed to evaluate protein expression in fisetin-treated and control cells, and Baculoviral IAP repeat-containing protein 8 (BIRC8) and apoptosis regulator Bcl-W (Bcl2L2) were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Their expression was confirmed by western blotting. The anticancer effect of fisetin in Huh-7 cells may result from the down regulation of BIRC8 and the Bcl2L2. [Copyright &y& Elsevier]

Abstract: The antiapoptotic BCL-2 family protein BCL-W is often overexpressed in colorectal carcinoma (CRC) where it correlates with advanced stage and expression of p53. In this work we have analysed the Bcl-w promoter to identify potential regulators of BCL-W expression in CRC cells. The Bcl-w promoter was highly active in cell lines derived from CRC as well as other cancer types. Although expression of p53 and BCL-W correlate in CRC, overexpression of wild type or mutant p53 did not significantly alter Bcl-w promoter activity, and deletion of endogenous p53 did not alter the expression of Bcl-w RNA in HCT116 cells. Promoter deletion analysis lead to the identification of a potential binding site for TCF/LEF factors, obligate binding partners for β-catenin, a downstream target of the WNT signalling pathway. TCF4 and β-catenin interacted with the Bcl-w promoter in intact HCT116 cells and mutation of this site significantly decreased promoter activity. The activity of the Bcl-w promoter was increased or decreased, respectively, by overexpression of β-catenin or dominant negative TCF4. β-catenin is activated in the majority of CRC and these results suggest that BCL-W may function as a downstream effector of inappropriate WNT/β-catenin signalling. [Copyright &y& Elsevier]

One of the most effective ways to combat different types of cancer is through early diagnosis and administration of effective treatment, followed by efficient monitoring that will allow physicians to detect relapsing disease and treat it at the earliest possible time. Apoptosis, a normal physiological form of cell death, is critically involved in the regulation of cellular homeostasis. Dysregulation of programmed cell death mechanisms plays an important role in the pathogenesis and progression of cancer as well as in the responses of tumours to therapeutic interventions. Many members of the BCL2 (B-cell CLL/lymphoma 2; Bcl-2) family of apoptosis-related genes have been found to be differentially expressed in various malignancies, and some are useful prognostic cancer biomarkers. We have recently cloned a new member of this family, BCL2L12 , which was found to be differentially expressed in many tumours. Most of the BCL2 family genes have been found to play a central regulatory role in apoptosis induction. Results have made it clear that a number of coordinating alterations in the BCL2 family of genes must occur to inhibit apoptosis and provoke carcinogenesis in a wide variety of cancers. However, more research is required to increase our understanding of the extent to which and the mechanisms by which they are involved in cancer development, providing the basis for earlier and more accurate cancer diagnosis, prognosis and therapeutic intervention that targets the apoptosis pathways. In the present review, we describe current knowledge of the function and molecular characteristics of a series of classic but also newly discovered genes of the BCL2 family as well as their implications in cancer development, prognosis and treatment. [ABSTRACT FROM AUTHOR]

Many long non-coding RNAs (lncRNAs) can exert crucial roles in the pathogenesis of cataract, including lncRNA KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1). We aimed to further elucidate the biological role and regulatory molecular mechanism of KCNQ1OT1 in cataract. The expression of KCNQ1OT1 and miR-223-3p and BCL2 like 2 (BCL2L2) was examined by qRT-PCR. Cataract cell model was constructed by treatment with hydrogen peroxide (H 2 O 2) in lens epithelial cells (SRA01/04). SRA01/04 cell viability and cell apoptosis were tested using CCK-8 assay and flow cytometry, respectively. Western blot (WB) was performed to measure the levels of apoptosis-related proteins and BCL2L2 protein. The oxidative stress factors were analyzed by corresponding kits. The interaction between miR-223-3p and KCNQ1OT1 or BCL2L2 was validated by dual-luciferase reporter and RNA Immunoprecipitation (RIP) assays. We found that KCNQ1OT1 was upregulated in cataract anterior lens capsule samples and H 2 O 2 -induced SRA01/04 cells. Knockdown of KCNQ1OT1 suppressed H 2 O 2 -induced SRA01/04 cell apoptosis and oxidative stress. KCNQ1OT1 acted as a sponge of miR-223-3p. Inhibition of miR-223-3p could abate the function of KCNQ1OT1 silence in H 2 O 2 -treated SRA01/04 cells. Additionally, BCL2L2 was a direct target of miR-223-3p, and miR-223-3p weakened H 2 O 2 -induced SRA01/04 cell apoptosis and oxidative stress by targeting BCL2L2. Collectively, the data suggest a role for the KCNQ1OT1/miR-223-3p/BCL2L2 axis in cataract formation but the data was generated using an epithelial cell line. • KCNQ1OT1 was upregulated in cataract. • KCNQ1OT1 knockdown alleviated H 2 O 2 -induced apoptosis of SRA01/04 cells by sponging miR-223-3p. • MiR-223-3p attenuated H 2 O 2 -induced apoptosis of SRA01/04 cells by targeting BCL2L2. [ABSTRACT FROM AUTHOR]

Objective: MiR-15 is involved in modulating many cellular processes that are normally destroyed in disease. The regulation of the cell cycle, angiogenesis, and responses to cellular stress are notable processes influenced by MiR-15. MiR-15 plays a critical role in the model of inflammation-related angiogenesis defects. However, its role in rheumatoid arthritis (RA) remains unclear. In this paper, we questioned the potential effects miR-15 has on RA., Methods: Synthetic mimics of miR-15 were transfected into human RA fibroblast-like synoviocyte (FLS) cell line MH7A cells. Cell proliferation was detected by a Cell Counting Kit-8 (CCK-8) assay. Cell apoptosis was determined by flow cytometry. MH7A cells, stimulated by interleukin-1β (IL-1β), were transfected with miR-15 mimics and inhibitors. Western blot and quantitative real time (qRT)-PCR were used to detect gene and protein expressions. Bioinformatic software analysis and luciferase reporter assays were used to predict and validate miR-15 targets., Results: We found that the miR-15 experienced down-regulated expression in MH7A cells, after they were stimulated with IL-1β. MiR-15 inhibited inflammatory cell proliferation and promoted inflammatory cell apoptosis. BCL2L2 was identified as a target of miR-15 by luciferase assay. Additionally, NF-κB signaling pathway related proteins were inhibited by miR-15 overexpression., Conclusion: These findings contribute to an understanding of the impact that miR-15 possesses on inflammatory injury, and provide a basis for RA treatment., (© 2019 by the Association of Clinical Scientists, Inc.)

Over 95% of human genes are alternatively spliced, expressing splice isoforms that often exhibit antagonistic functions. We describe genes whose alternative splicing has been linked to prostate cancer; namely VEGFA, KLF6, BCL2L2, ERG, and AR. We discuss opportunities to develop novel therapies that target specific splice isoforms, or that target the machinery of splicing. Therapeutic approaches include the development of small molecule inhibitors of splice factor kinases, splice isoform specific siRNAs, and splice switching oligonucleotides. [ABSTRACT FROM AUTHOR]